Recently, it has been shown that a variety of drugs require carrier-mediated transport (e.g., the human dipeptide transporter or hpepT1) in order to cross the cell membrane and be absorbed into the body. Although the use of membrane transporters to enhance drug absorption is promising, detailed knowledge necessary to apply this approach is not presently available. Thus, the long-term goal of this research program is the development of in vitro tissue culture systems for screening of transporter-dependent drugs or prodrugs. Reproducibility, cost effectiveness, and adaptability to different transporter delivery mechanisms and drugs would characterize the ideal system. The central hypothesis of the research proposed is that hpepT1-dependent regulation of carrier-mediated absorption can be increased in cell lines to allow the development of in vitro assays to predict intestinal permeability of peptidometic drugs. The specific aims will utilize a human cell culture system in which we will increase expression of intestinal hpepT1 transporter to more closely approximate the in vivo intestinal expression of the transporter. We will develop an in vitro competitive inhibition assay for peptidometic compounds. We will validate this screening model by correlation of the rank-order of the paired measurements of competitive inhibition (K1) in vitro and rat intestinal permeability in situ (Peff) for a series of 20 peptidometic compounds. PROPOSED COMMERCIAL APPLICATIONS: The successful development of in vitro competitive inhibition assay for screening peptidomimetic compounds will provide a new tool for development of new drugs that require membrane transporters for absorption. This screening system is expected to provide the basis for future intestinal transporter screens to aid drug development.